M. Andrei et al. / European Journal of Medicinal Chemistry 46 (2011) 5935e5940
5939
A solution of the thioic acid 4a (0.170 g, 0.3 mmol) and NH4F
4.8. (RP)-8-(2-furyl)-N6,N6-dimethyladenosine-30,50-cyclic phosph
(0.075 g, 2 mmol) in DMF (1.5 mL) was stirred at room temperature
for 5 d. The reaction mixture was filtered, n-Bu3N (0.111 g,
0.6 mmol) was added to the clear filtrate and the solvent was
removed at reduced pressure. The residue was washed with hexane
to remove the excess of n-Bu3N and the residual material subjected
to flash chromatography on silica gel using CH2Cl2:MeOH: n-Bu3N
100:10:1. The ammonium salt, which contained some residual
n-Bu3N, was further purified by dissolution in CH2Cl2 and repreci-
pitation by hexane addition; yield 0.105 g (37%) of a white solid.
HRMS (electrospray): Found for M-NHBu3: 438.0642. Calc. for
C28H45N76O6PSH-N(C4H9)3: 438.0652; 31P (121 MHz, MeOH-d4,):
orothioic acid sodium salt (6a)
(RP)-8-(2-Furyl)-N6,N6-dimethyladenosine-30,50-cyclic
phos-
phorothioic acid tri-n-butylammonium salt (5a) (100 mg,
0.16 mmol) was dissolved in methanolic 0.1 M NaOH (1.7 mL). The
sodium salt was precipitated by addition of diethyl ether and
collected by filtration; yield 66 mg (89%) of a white solid material.
HRMS (electrospray): Found for M-Na: 438.0645. Calc. for
C16H18N5O6PS: 438.0652; 31P (121 MHz, MeOH-d4):
d
57.9; 1H NMR
3.05 (bs, 6H, N(CH3)2), 3.93e4.24 (m, 3H),
(200 MHz, MeOH-d4):
d
4.50e4.54 (m, 2H), 5.06 (d,1H, J ¼ 5.3 Hz), 5.36e5.40 (m,1H), 6.14 (s,
d
57.9; 1H NMR (200 MHz, MeOH-d4):
d
0.93 (t, 9H, J ¼ 7.3 Hz,
1H), 6.53e6.56 (m,1H), 7.05e7.08 (m,1H), 7.58e7.61 (m,1H), 8.31 (s,
3 ꢂ CH3), 1.29e1.41 (m, 6H, 3 ꢂ CH2), 1.55e1.65 (m, 6H, 3 ꢂ CH2),
2.98e3.04 (m, 6H, 3 ꢂ CH2), 3.03 (bs, 6H, N(CH3)2), 3.92e4.26 (m,
3H), 4.50e4.54 (m, 2H), 5.02 (d, 1H, J ¼ 5.3 Hz), 5.39e5.43 (m, 1H),
6.13 (s, 1H), 6.53e6.56 (m, 1H), 7.04e7.07 (m, 1H), 7.58e7.61 (m,
1H, H-2); 13C NMR (75 MHz, MeOH-d4):
d 68.5, 78.2, 73.2, 77.6, 94.2,
113.1, 115.1, 120.8, 142.9, 144.7, 146.6, 149.8, 153.9, 156.3. No 13C-
signal for N(CH3)2 was seen.
1H), 8.30 (s,1H, H-2); 13C NMR (75 Mz, MeOH-d4):
d 13.9, 20.9, 26.8,
4.9. (RP)-8-(2-Furyl)-N6-methyladenosine-30,50-cyclic phosphoro
thioic acid sodium salt (6b)
53.9, 68.5, 72.8, 73.2, 77.7, 94.2, 113.1, 115.1, 120.8, 142.9, 144.7, 146.6,
149.8, 153.9, 156.4. No 13C-signal was observed for N(CH3)2.
Compound (6b) was prepared from the tri-n-butylammonium
salt (5b) (0.16 mmol) with methanolic 1 M NaOH (1.7 mL) as above;
yield 90% of a white solid material. HRMS (electrospray): Found for
M-Na: 424.0473. Calc. for C15H15N5O6PS: 424.0471; 31P (121 MHz,
4.6. (RP)-8-(2-Furyl)-N6-methyladenosine-30,50-cyclic phosphoro
thioic acid tri-n-butylammonium salt (5b)
MeOH-d4);
d d 3.14 (bs, 3H),
57.9; 1H NMR (200 MHz, MeOH-d4):
Compound (5b) was prepared as above from (SP)-20O-(tert-
butyldimethylsilyl)-8-(2-furyl)-N6-methyladenosine-30,50-cyclic
N-benzylphosphoramidate (3b) (0.637 mmol) and CS2 (1.9 mmol).
The initial product (RP)-20O-(tert-butyldimethylsilyl)-8-(2-furyl)-
N6-methyladenosine-30,50-cyclic phosphorothioic acid (4b) was
desilylated with NH4F (4.05 mmol) in DMF (3 mL) and purified as
its n-Bu3N salt by flash chromatography on silica gel using
CH2Cl2:MeOH: n-Bu3N 100:5:1; yield 0.140 g (36%) of a white
solid. HRMS (electrospray): Found for M-NHBu3 424.0475. Calc.
for C27H43N6O6PSH-N(C4H9)3: 424.0471. 31P (121 MHz, MeOH-d4):
4.29e4.42 (m, 3H), 5.18 (d, 1H, J ¼ 5.3 Hz,), 5.49e5.57 (m, 1H), 6.28
(s, 1H), 6.73e6.75 (m, 1H), 7.21e7.22 (m, 1H), 7.86e7.87 (m, 1H),
8.31 (s, 1H, H-2); 13C NMR (75 Mz, MeOH-d4,):
d 68.4, 72.8, 73.2,
77.6, 94.2, 113.1, 115.1, 120.8, 142.9, 144.7, 146.6, 149.9, 154.2, 156.4.
No 13C-signal for NHCH3 was seen.
4.10. (RP)-N6-Benzyl-8-(2-furyl)adenosine-30,50-cyclic phosphoro
thioic acid sodium salt (6c)
d
57.9; 1H NMR (200 MHz, MeOH-d4):
d
0.95e1.02 (t, 9H, J ¼ 7.3 Hz,
Compound (6c) was prepared as above from the tri-n-buty-
lammonium salt (5c) (0.14 mmol) in methanolic 0.1 M NaOH
(1.5 mL); yield 90% of a white, solid material. HRMS (electrospray):
Found for M-Na: 500.0799. Calc. for C21H19N5O6PS: 500.0778; 31P
3 ꢂ CH3), 1.32e1.47 (m, 6H, 3 ꢂ CH2), 1.61e1.73 (m, 6H, 3 ꢂ CH2),
3.08e3.14 (m, 6H, 3 ꢂ CH2), 3.17 (bs, 3H), 4.27e4.34 (m, 3H), 5.13
(d, 1H, J ¼ 5.3 Hz), 5.39e5.44 (m, 1H), 6.24 (s, 1H), 6.68e6.70 (m,
1H), 7.16e7.18 (m, 1H), 7.81e7.82 (m, 1H), 8.26 (s, 1H, H-2); 13C
(121 MHz, MeOH-d4):
d
57.9; 1H NMR (200 MHz, MeOH-d4):
NMR (MeOH-d4, 75 Mz):
d 13.9, 20.9, 26.8, 53.9, 68.5, 72.8, 73.2,
d
4.20e4.35 (m, 3H), 4.77 (bs, 2H), 5.18 (d, 1H, J ¼ 5.3 Hz), 5.49e5.53
77.7, 94.2, 113.1, 115.1, 120.8, 143.0, 144.7, 146.6, 149.8, 153.9, 156.3.
(m, 1H), 6.24 (s, 1H), 6.64e6.66 (m, 1H), 7.13e7.14 (m, 1H), 7.15e7.34
No 13C-signal for NHCH3 was seen.
(m, 5H), 7.76e7.77 (m, 1H), 8.25 (s, 1H, H-2); 13C NMR (75 Mz,
MeOH-d4):
d 68.5, 72.8, 73.2, 77.6, 94.2, 113.1, 115.2, 120.5, 128.2,
128.5, 129.5, 140.2, 143.1, 144.7, 146.6, 150.9, 154.2, 155.6. No 13C-
4.7. (RP)-N6-Benzyl-8-(2-furyl) adenosine-30,50-cyclic phosphoro
thioic acid tri-n-butylammonium salt (5c)
signal for NHCH2Ph was seen.
Compound (5c) was prepared as above from (SP)-N6-benzyl-20O-
(tert-butyldimethylsilyl)-8-(2-furyl)adenosine-30,50-cyclic N-ben-
zylphosphoramidate (3c) (0.51 mmol) and CS2 (1.53 mmol). The
initial product (RP)-N6-benzyl-2’O-(tert-butyldimethylsilyl)-8-(2-
furyl) adenosine-30,50 cyclic phosphorothioic acid (4c) was desily-
lated by NH4F (2.7 mmol) in DMF (2 mL) and purified as its n-Bu3N
salt by flash chromatography on silica gel using CH2Cl2:MeOH:
n-Bu3N 100:5:1; yield 0.135 g (39%) of a white solid. HRMS (elec-
trospray): Found for M-NHBu3 500.0799. Calc. for C33H47N6O6PSH-
4.11. (Rp)-6-N-Butyryl-20-O-(tert-Butyldimethylsilyl)-8-(2-furyl)
adenosine ꢁ30,50-cyclic phosphorothioic acid tri-n-butylammonium
salt (8b)
A 1.8 M solution of LDA in THF/heptane/ethylbenzene (1.27 mL,
2.3 mmol) was added to a solution of (Sp)-20-O-tert-butyldime-
thylsilyl)-8-(2-furyl)adenosine-30,50-cyclic
N-benzylphosphor-
amidate (1) (1.22 g, 2.0 mmol) in THF (25 mL) under argon
at ꢁ78 ꢀC. The mixture was stirred for 10 min at this temperature
before CS2 (0.362 mL, 6 mmol) was added. The cooling bath was
removed after 10 min. The mixture was stirred for 3 h at room
temperature before the solution was concentrated under reduced
pressure to about 10 mL when hexane was added until precipita-
tion was completed (ca 60 mL). The precipitate was collected, dried,
and suspended in water (25 mL). Subsequently, 1 M HBr was added
to pH 1e2 when the phosphorothioic acid 7 was precipitated,
filtered off and dried under reduced pressure; yield 0.945 g (90%) of
a light tan solid.
N(C4H9)3: 500.0785. 31P (121 MHz, MeOH-d4,):
(200 MHz, MeOH-d4):
d
57.9; 1H NMR
d
0.96e1.01 (t, 9H, J ¼ 7.3 Hz,), 1.37e1.45 (m,
6H, 3 CH2), 1.63e1.671 (m, 6H, 3 e CH2), 3.09e3.15 (m, 6H, 3 CH2),
4.20e4.35 (m, 3H), 4.79 (bs, 2H), 5.15 (d, 1H, J ¼ 5.3 Hz), 5.49e5.53
(m,1H), 6.25 (s, 1H), 6.66e6.68 (m,1H), 7.16e7.17 (m,1H), 7.21e7.38
(m, 5H), 7.79e7.80 (m, 1H), 8.26 (s, 1H, H-2); 13C NMR (75 Mz,
MeOH-d4,):
d 13.9, 20.9, 26.8, 53.9, 68.5, 72.8, 73.2, 77.6, 94.2, 113.1,
115.2, 120.6, 128.2, 128.5, 129.5, 140.2, 143.1, 144.7, 146.7, 150.9,
154.1, 155.6. No 13C-signal for NHCH2Ph was seen.